• Archives of Pharmacal Research
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• 제27권3호
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• pp.324-333
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• 2004

To determine whether Insulin-like growth factor (IGF-I) treatment represents a potential means of enhancing the survival of cardiac muscle cells from adriamycin (ADR)-induced cell death, the present study examined the ability of IGF-I to prevent cell death. The study was performed utilising the embryonic, rat, cardiac muscle cell line, H9C2. Incubating cardiac muscle cells in the presence of adriamycin increased cell death, as determined by MTT assay and annexin V-positive cell number. The addition of 100 ng/mL IGF-I, in the presence of adriamycin, decreased apoptosis. The effect of IGF-I on phosphorylation of PI, a substrate of phosphatidylinositol 3-kinase (PI 3-kinase) or protein kinase B (AKT), was also examined in H9C2 cardiac muscle cells. IGF-I increased the phosphorylation of ERK 1 and 2 and $PKC{\;}{\zeta}{\;}kinase$. The use of inhibitors of PI 3-kinase (LY 294002), in the cell death assay, demonstrated partial abrogation of the protective effect of IGF-I. The MEK1 inhibitor-PD098059 and the PKC inhibitor-chelerythrine exhibited no effect on IGF-1-induced cell protection. In the regulatory subunit of PI3K-p85- dominant, negative plasmid-transfected cells, the IGF-1-induced protective effect was reversed. This data demonstrates that IGF-I protects cardiac muscle cells from ADR-induced cell death. Although IGF-I activates several signaling pathways that contribute to its protective effect in other cell types, only activation of PI 3-kinase contributes to this effect in H9C2 cardiac muscle cells.

• The Korean Journal of Physiology and Pharmacology
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• 제5권1호
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• pp.1-8
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• 2001

Hyperpolarization of arterial smooth muscle by acetylcholine is considered to be produced by the release of an unidentified chemical substance, an endothelium-derived hyperpolarizing factor (EDHF). Several chemicals have been proposed as the candidate for EDHF. However, none of them fulfil completely the nature and property of EDHF. Ultrastructural observation with electron microscope reveals that in some arteries, gap junctions are formed between endothelial and smooth muscle cells. In small arterioles, injection of gap junction permeable dyes into an endothelial cell results in a distribution of the dye to surrounding cells including smooth muscle cells. These observations allow the speculation that myoendothelial gap junctions may have a functional significance. Simultaneous measurement of the electrical responses in both endothelial and smooth muscle cells using the double patch clamp method demonstrates that these two cell types are indeed electrically coupled, indicating that they behave as a functional syncytium. The EDHF-induced hyperpolarization is produced by an activation of $Ca^{2+}-sensitive\;K^+-channels$ that are inhibited by charybdotoxin and apamin. Agonists that release EDHF increase $[Ca^{2+}]_i$ in endothelial cells but not in smooth muscle cells. Inhibition of gap junctions with chemical agents abolishes the agonist-induced hyperpolarization in smooth muscle cells but not in endothelial cells. All these observations can be explained if EDHF is an electrotonic signal propagating from endothelium to smooth muscle cells through gap junctions.

• Molecules and Cells
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• 제21권1호
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• pp.42-51
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• 2006

We investigated the mechanism of contraction induced by S1P in esophageal smooth muscle cells. Western blot analysis demonstrated that $S1P_1$, $S1P_2$, $S1P_3$, and $S1P_5$ receptors existed in the cat esophagus. Only penetration of EDG-5 ($S1P_2$) antibody into permeabilized cells inhibited S1P-induced contraction. Pertussis toxin (PTX) also inhibited contraction, suggesting that it was mediated by $S1P_2$ receptors coupled to a PTXsensitive $G_i$ protein. Specific antibodies to $G_{i2}$, $G_q$ and $G_{\beta}$ inhibited contraction, implying that the S1P-induced contraction depends on PTX-insensitive $G_q$ and $G_{\beta}$ dimers as well as the PTX-sensitive $G_{i2}$. Contraction was not affected by the phospholipase $A_2$ inhibitor DEDA, or the PLD inhibitor ${\rho}$-chloromercuribenzoate, but it was abolished by the PLC inhibitor U73122. Incubation of permeabilized cells with $PLC{\beta}3$ antibody also inhibited contraction. Contraction involved the activation of a PKC pathway since it was affected by GF109203X and chelerythrine. Since $PKC{\varepsilon}$ antibody inhibited contraction, $PKC{\varepsilon}$ may be required. Preincubation of the muscle cells with the MEK inhibitor PD98059 blocked S1P-induced contraction, but the p38 MAP kinase inhibitor SB202190 did not. In addition, co-treatment of cells with GF 109203X and PD98059 did not have a synergistic effect, suggesting that these two kinases are involved in the same signaling pathway. Our data suggest that S1P-induced contraction in esophageal smooth muscle cells is mediated by $S1P_2$ receptors coupled to PTX-sensitive $G_{i2}$ proteins, and PTX-insensitive $G_q$ and $G_{\beta}$ proteins, and that the resulting activation of the $PLC{\beta}3$ and $PKC{\varepsilon}$ pathway leads to activation of a p44/p42 MAPK pathway.

• 대한한의학방제학회지
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• 제29권1호
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• pp.45-55
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• 2021

Objectives : This study was conducted to evaluate the anti-atrophic effect of polycan in dexamethasone-induced skeletal muscle atrophy in vitro model. Methods : C2C12 myoblast were differentiated into myotube by 2% horese serum medium for 6 days, and then treated polycan extract at different concentrations for 24h. The effect of dexamethasone on the induction of muscle atrophy and expression of atrophy-related genes in differentiated C2C12 myotubes using a GSH, ROS, real-time PCR, western blots analysis. Results : The results showed that Treatment with polycan (100 and 200 ㎍/㎖) noncytotoxic levels on both myoblast and myotube. Polycan decreased the ROS level overproduced with dexamethasone and improved the depletion of GSH level. Dexamethasone showed a decrease in myotube diameter, which was associated with up-regulation muscle-specific ubiquitin ligases markers, such as atrogin-1, FoxO3, myostatin and muscle RING finger-1 (MuRF1), and down-regulation of myogenin, MEF2, Myogenic regulatory factor 5, 6 and MyoD. The results showed that polycan treatment significantly dose-dependently inhibited it. Furthermore, decreased expressions of PI3K/Akt signal pathway by dexamethasone were reversed by treatment with polycan. Conclusions : Thus, polycan suppresses dexamethasone induced muscle atrophy in C2C12 myotube in vitro model through activation of PI3K/Akt pathway and protective effect of improve skeletal muscle function.

• BMB Reports
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• 제50권12호
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• pp.628-633
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• 2017

Gastrin-releasing peptide (GRP) has been reported to be implicated in the pathogenesis of inflammatory disorders. The migration and proliferation of vascular smooth muscle cells (VSMCs) are key components of vascular inflammation that leads to the development of atherosclerosis. The present study aimed to investigate the molecular effect of GRP on VSMC proliferation and migration. We report that GRP significantly enhanced the proliferation and migration of rat VSMCs. GRP increased mRNA and protein expression of matrix metalloproteinase-2 and -9 (MMP-2/9) in VSMCs. The induction of MMP-2/9 by GRP was regulated by the activation of the signal transducer and activator of transcription-3 (STAT3). In addition, STAT3-knockdown of VSMCs by siRNA or blockade of the GRP receptor inhibited GRP-induced migration of VSMCs. Taken together, our findings indicate that GRP promotes the migration of VSMCs through upregulation of MMP-2/9 via STAT3 activation.

• Molecules and Cells
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• 제20권3호
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• pp.429-434
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• 2005

Lysophosphatidylcholine (lysoPC) induces vascular smooth muscle cell (VSMC) proliferation and migration, which has been proposed to initiate the intimal thickening in coronary atherosclerotic lesions. Berberine is an alkaloid in Berberis aquifolium and many other plants. Recently, it has been shown to have beneficial effects on the cardiovascular system, such as anti-hyperglycemic and cholesterol-lowering activity. In this study, we investigated its effects on lysoPC-induced VSMC proliferation and migration. Berberine inhibited lysoPC-induced DNA synthesis and cell proliferation in VSMCs, as well as migration of the lysoPC-stimulated VSMCs. It also inhibited the activation of extracellular signal-regulated kinases (ERKs) and reduced transcription factor AP-1 activity and the lysoPC-induced increases in intracellular reactive oxygen species (ROS). These results indicate that the inhibitory effects of berberine on lysoPC-stimulated VSMC proliferation and migration are attributable to inhibition of ROS generation and hence of activation of the ERK1/2 pathway. This suggests that berberine has potential in the prevention of atherosclerosis and restenosis.

• 생명과학회지
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• 제21권1호
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• pp.36-43
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• 2011

Glycate화된 단백질이 혈관질환의 발생에 관여하는지 알아보기 위하여 glycated albumin (GA)이 혈관평활근 세포에서 인터루킨-6 발현에 영향을 주는지 조사하고 또한 그 기전을 구명하였다. GA에 노출된 혈관평활근세포에서 인터루킨-6 transcript가 증가하고, 인터루킨-6 단백질의 분비가 증가하고, 또한 인터루킨-6 유전자의 promoter가 활성화되었다. GA에 의한 인터루킨-6 유전자의 promoter 활성화는 dominant negative 형태의 Toll-like receptor (TLR)-4와 myeloid differentiation factor 88 (Myd88)에 의하여 크게 감소되었지만, dominant negative 형태의 TLR-2와 TIR-domain-containing adapter-inducing interferon-$\beta$ (TRIF)의 영향을 받지 않았다. 그리고 Extrcellular signal-related kinase (ERK) 억제 물질들은 GA에 의한 인터루킨-6의 분비 및 인터루킨-6 유전자 promoter 활성화를 억제하였다. 그리고 인터루킨-6 유전자의 promoter의 NF-${\kappa}B$-binding sequence에 변이는 GA에 의한 인터루킨-6 유전자의 promoter 활성화 억제하였다. 이러한 결과는 혈관평활근세포에서 GA에 의한 인터루킨-6 유전자 활성화에 TLR-4와 ERK 및 NF-${\kappa}B$가 관여함을 의미한다.

• Archives of Pharmacal Research
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• 제29권1호
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• pp.67-72
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• 2006

The abnormal proliferation of aortic vascular smooth muscle cells (VSMCs) plays a central role in the pathogenesis of atherosclerosis and restenosis after angioplasty and possibly also in the development of hypertension. The present study was designed to examine the inhibitory effects and the mechanism of luteolin 7-glucoside (L7G) on the platelet-derived growth factor (PDGF)-BB-induced proliferation of VSMCs. L7G significantly inhibited the PDGF-BB-induced proliferation and the DNA synthesis of the VSMCs in a concentration-dependent manner. Pre-incubation of the VSMCs with L7G significantly inhibited the PDGF-BB-induced extracellular signal-regulated kinase 1/2 (ERK1/2), Akt and the phospholipase C $(PLC)-{\gamma}1$ activation. However, L7G had almost no affect on the phosphorylation of $PDGF-{\beta}$ receptor tyrosine kinase, which was induced by PDGF-BB. These results suggest that L7G inhibits the PDGF-BB-induced proliferation of VSMCs via the blocking of $(PLC)-{\gamma}1$, Akt, and ERK1/2 phosphorylation.

• BMB Reports
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• 제46권7호
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• pp.352-357
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• 2013

Atherosclerosis, which manifests as acute coronary syndrome, stroke, and peripheral arterial diseases, is a chronic inflammatory disease of the arterial wall. Prunella vulgaris, a perennial herb with a worldwide distribution, has been used as a traditional medicine in inflammatory disease. Here, we investigated the effects of P. vulgaris ethanol extract on TNF-${\alpha}$-induced inflammatory responses in human aortic smooth muscle cells (HASMCs). We found that P. vulgaris ethanol extract inhibited adhesion of monocyte/macrophage-like THP-1 cells to activated HASMCs. It also decreased expression of intercellular adhesion molecule-1, vascular cell adhesion molecule-1, E-selectin and ROS, No production in TNF-${\alpha}$-induced HASMCs and reduced NF-${\kappa}B$ activation. Furthermore, P. vulgaris extract suppressed TNF-${\alpha}$-induced phosphorylation of p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinase (ERK). These results demonstrate that P. vulgaris possesses anti-inflammatory properties and can regulate TNF-${\alpha}$-induced expression of adhesion molecules by inhibiting the p38 MAPK/ERK signaling pathway.

• Nutritional Sciences
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• 제7권1호
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• pp.23-30
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• 2004

As a central component of a novel protein kinase cascade, the activation of the mitogen-activated protein (MAP) kinase cascade has attracted considerable attention. We sought to determine the effect of exercise and diet on the activation of the extracellular-signal regulated protein kinase (ERK) 1/2 and the p38 MAP kinase pathways in rat soleus muscle. Forty-eight Sprague-Dawley rats were assigned to one of two dietary conditions: high-carbohydrate (CHO) or high-fat (FAT). Animals having each dietary condition were further divided into one of three subgroups: a sedentary control group that did not exercise (NT), a group that performed 8 weeks of treadmill running and was sacrificed 48 h after their final treadmill run (CE), and a group that was sacrificed immediately after their final routine exercise training (AE). A high-fat diet did not have any significant effect on phosphorylated and total forms of ERK 1/2 or p38 MAP kinase. In chronically trained muscle that was taken 48 h after the last training, phosphorylated ERK 1/2 significantly increased only in the FAT but not in the CHO groups. In the case of total ERK 1/2, it increased significantly for both groups. In contrast, both phosphorylated and total forms of p38 MAP kinase decreased markedly compared to sedentary muscle. In muscle that was taken immediately after a last bout of exercise, phosphorylated ERK 1/2 increased in both groups but statistical significance was seen only in the CHO group. Total ERK 1/2 in acutely stimulated muscle increased only in the CHO-AE group even though the degree was much lower than the phosphorylated status. Muscle that was taken immediately after the routine training increased in phosphorylation status of p38 MAP kinase for both dietary conditions. However, statistical significance was seen only in the CHO group owing to a large variation with FAT. In conclusion, a high-fat diet per se did not have any notable effect versus a high-carbohydrate diet on MAP kinase pathways. However, when diet (either CHO or FAT) was combined with exercise and/or training, there was differentiated protein expression in MAP kinase pathways. This indicates MAP kinase pathways have diverse control mechanisms in slow-twitch fibers.